Theses and dissertations (Applied Sciences)
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Item Electrochemical aptasensor for the detection of mycotoxins in food samples by experimental and computational methods(2021) Kunene, Kwanele; Bisetty, Krishna; Kanchi, Suvardhan; Sabela, Myalowenkosi InnocentMycotoxins are secondary metabolites of fungi that are present in various foodstuff and feed commodities. A large number of mycotoxins exist, however only a limited number represent a major damages and toxic properties. Amongst them, the aflatoxins and ochratoxins are deemed to be the most poisonous and extensively circulated in the world and then, represent a real hazard to both human and animal. Depending on several factors like the consumption levels, exposure time, mechanisms of action, digestion and defense mechanisms, mycotoxins stimulate a wide spectrum of toxicological effects leading to both acute and chronic diseases, liver and kidney failure, skin rash, cancer, immune suppression, birth defects or even death. To address the harmful impact of mycotoxin contaminants in food and feed, health authorities in various countries world-wide have established guidelines in order to protect human and animal from the possible damages instigated by these toxins. Authorities such as the European Commission, US Food and Drug Administration (FDA), World Health Organization (WHO) and the Food and Agriculture Organization of the United Nations (FAO) set up maximum level regulations for main mycotoxins in foods and feeds. To accomplish the expectations of these regulation levels, there is a great need for the development and validation of modern, uncomplicated, rapid, and detailed methodologies for the detection of toxins. In this study, various approaches for the rapid, inexpensive and ultrasensitive biosensors for the detection of two major mycotoxins were developed. The electrochemical-based aptasensor and immunosensor were developed for the determination of aflatoxin B1 (AFB1) and ochratoxin A (OTA) in different food products. The fabricated biosensors demonstrated good practical analytical feasibility for mycotoxins detection in real samples such as WeetBix, yoghurt, coffee and in wine samples with excellent recoveries and RSD values. To avoid fouling on the sensor surface by the constituents present in real samples, the carbon screen printed electrode (C-SPE) and carbon felt electrode (CFE) surfaces were modified with different nanomaterials such as silver nanoparticle (AgNPs), palladium nanoparticles (PdNPs), palladium doped boron nitride (PdNPs-BN) and titanium nanoparticles doped with boron nitride BN-TiO2. In addition, the aptamers and antibodies were immobilized on the modified electrode in order to enhance the selectivity of the sensor towards the detection of OTA and AFB1. The electrochemical aptasensor for OTA permitted for highly sensitive detection in Weet-Bix with a wide linear range (0.002 - 0.016 mg L-1) and limit of detection of 7×10-4 mg L-1. It is worth prominence that it is the first time that carbon screen printed electrode (C-SPE) modified with AgNPs was used, opening new pathways for highly precise analysis. Experimental results were further supported computationally for a better understanding of the interaction between the aptamer and the analytes. Computational results were in good agreement with experimental results. The same procedure was also established in voltammetric detection of AFB1 using CFE modified with BN-TiO2 (CF/BN-TiO2). A wide concentration range of 2.5 - 20 ng mL-1 with an excellent LOD of 0.002 ng mL-1 for AFB1 was obtained. For the case study of wine samples tested for AFB1 detection, a simple but very effective pretreatment method was effectively applied. The addition of acetonitrile to the wine reduces the non-specific interactions that might be accountable for inactivation of antibody and blocking of the sensor surface. Furthermore, the PdNPs-BN enhanced the electrical signal and the sensor sensitivity. Attained results allowed for AFB1 detection at concentrations range from 1.0 - 10 ng mL-1 with limit of detection of 0.832 ng mL-1 . In the case study of the electrochemical immunosensor for the detection of OTA in coffee, a linear detection range of 0.5 - 20 ng mL-1 was achieved with LOD of 0.096 ng mL-1 . The fabricated aptasensors and immunosensors in this study combines the most desirable characteristics of a good biosensor such as high sensitivity, inexpensive, rapid, and simple but portable method make proposed approaches an important and very promising tools for extensive biosensing applications.Item Novel sensors for detection of anti-asthma drugs in pharmaceutical formulations(2021) Kilele, Chepkoech Joan; Redhi, Gan G.; Chokkareddy, RajasekharThe development of an effective technique for detecting anti-asthma drugs with adverse effects in the human body at high dosages remains a serious challenge. For this purpose, herein, we report a novel electrochemical detection method based on nanocomposite modified glassy carbon electrode (GCE) nano-sensors. Cobalt ferrite (CoFe2O4), titanium oxide (TiO2) and zinc oxide (ZnO) nanoparticles (ZnONPs) were synthesized as well as their respective nanocomposites (CoFe2O4-MWCNTs-Lipase, TiO2-MWCNTs-IL, ZnONPs-MWCNTsCytochrome c and MWCNTs-Ionic liquid-L-lysine) were successfully prepared. Subsequently, their surface morphology, crystal structure, functional groups, thermal stability, surface area as well as pore size distribution of the synthesized nanoparticles and the nanocomposites aspects were analysed. The characterization analyses were undertaken using scanning electron microscopy (SEM), transmission electron microscopy (TEM), X-ray diffraction (XRD), Brunauer-Emmett Teller (BET) adsorption analyses, thermal gravimetric analysis (TGA) and Fourier-transform infrared spectroscopy (FTIR). In addition, electrochemical characterization of the fabricated electrodes were examined using electrochemical impedance spectroscopy (EIS) in order to ensure the material’s suitability in electro-catalytic sensing by investigating the electron transfer kinetics of the redox probe at the modified electrode-solution interface. The fabricated nanocomposite based sensors; ZnONPs-MWCNTs-Cyt c-GCE, CoFe2O4- MWCNTs-Lipase-GCE, MWCNTs-IL-L-lysine-GCE and TiO2-MWCNTs-IL-GCE were tested for electrochemical determination of theophylline, salbutamol, prednisolone and terbutaline, respectively, using cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The effect of varying scan rates, pH of the supporting electrolyte and accumulation time on the peak potential position and peak-currents, were studied with each electrode, and the optimum working parameters were selected. The ZnONPs-MWCNTs-Cyt c-GCE, CoFe2O4-MWCNTs-Lipase-GCE, MWCNTs-IL-L-lysine-GCE and TiO2-MWCNTs-IL-GCE sensors were found to exhibit excellent electro-catalytic activity towards theophylline, salbutamol, prednisolone and terbutaline. The synergy between these nanocomposite materials increased the electrochemical activity, the electron transfer rate and the electrode surface area, leading to high peak-current response for the determination of each drug. ZnONPs-MWCNTs-Cyt c-GCE sensor presented acceptable long-term stability, good reproducibility and repeatability. The limit of detection (LOD = 0.0012 µM) obtained was much lower than most of the given literature studies. Similarly, the CoFe2O4-MWCNTs-Lipase nanocomposite modified GCE for salbutamol (SAL) exhibited excellent stability, reproducibility and sensitivity with lowest detection limit (0.0240 µM) and limit of quantification (LOQ) of 0.0120 µM, over the concentration range of 0.05- 3.9 µM. In addition, the MWCNTs-IL-L-lysine-GCE exhibited outstanding electro-catalytic activity including excellent reproducibility, with LOD and LOQ of 0.0214 μM and 0.3016 μM, respectively, for prednisolone (PDN) detection. Moreover, at optimized electrochemical experimental conditions, the electrochemical performance of the constructed TiO2-MWCNTs-IL-GCE sensor for determination of terbutaline (TBS) was also evaluated. Stability, reproducibility as well as the detection limits were determined. A wide linear range of 0.5 µM to 3.0 µM as well as LOD and LOQ of 0.0162 µM and 0.2140 µM, respectively, were obtained and the results were compared with similarly reported electrodes for terbutaline detection. The selectivity of each sensor was also evaluated in presence of common interferences. There was no significant interference on the electrochemical peak-current response and peak position of each target analyte. The practical applicability of each fabricated sensor was investigated by the DPV method with respect to the detection of the four drugs under study in commercial pharmaceutical samples. In all cases, the sensors showed good percentage recovery performance in real dosage samples. From the experimental results, it is evident that the fabricated nanocomposite based sensors are ultra-sensitive for selective detection of the investigated four drugs in pharmaceutical formulations and other sample matrices.